Cytochrome P450 monooxygenases often determine the clearance of candidate drugs limiting their efficacy. These enzymes can also be sources for off-target toxicity. Our long term objective is to understand the structural determinants of substrate recognition by P450 enzymes. Collectively and individually the human drug metabolizing P450s can metabolize structurally diverse range of substrates, and this is likely to reflect in part the flexibility of each enzyme and structural adaptations for substrate binding. This underscores the necessity to determine multiple structures of individual enzymes in complex with structurally dissimilar drugs to understand the contribution of conformational flexibility to drug binding. Specific changes in the active site architectures of P450s 1B1, 1A2, 2D6, and 2C19 induced by interaction with chemically diverse substrates, inhibitors and activators, alone or in combination, will be identified to delineate the range of adaptive changes that can occur for each enzyme. Additionally, the active site topologies of P450s 2J2, 3A5 and 3A7 as well as representative family 4 enzymes will also be determined to identify structural characteristics that control the important functional contribution made by these enzymes to drug metabolism and the clearance of xenobiotics and excess endogenous compounds. Collectively, these studies will address significant gaps in our knowledge of P450 structure as it relates to function, and provide important information and tools for the prediction of drug metabolism. PUBLIC HEALTH RELEVANCE: These studies will provide new information to guide drug design in order to avoid limitations of efficacy and undesirable off-target effects resulting from cytochrome P450 mediated drug metabolism. This will benefit a wide range of therapeutic areas targeted for drug development.